Bacteriological analysis of the water in El Caño Martín Peña

Detection and quantification of Enteric Pathogenic Bacteria Salmonella and Shigella spp.

1 / march/ 2013

Mario R. BarreraGeorge M. RiveraPaola TabaroMentor: Prof. Mayra RolónUniversity of the Sacred Heart

Meet the Team

Mario R.Barrera

PaolaTabaro

George M. Rivera

Mentor:Prof: Mayra Rolón

Meet the Team

2012

2013

Caño Martín Peña

Is an invaluable natural habitat for the Metropolitan area of San Juan.

Caño Martín Peña

+ During the recurring years the mangroves and lagoons stretched of the Caño have been contaminated and stretched out into smaller measurements.

Caño Martín PeñaThe deterioration of this area has taken its toll on the residents lifestyles.

How to Improve !! The decontamination

of the lagoon that boards the community.

Rehabilitation of the estuary system.

To broaden the community’s knowledge of the microbial presence and the harms that can be encountered.

Our objectives

+ To detect the presence of pathogenic enteric bacteria such as Salmonella and Shigella SPP.

+ To acquire knowledge of the microbial presence found in the Caño Martín Peña.

+ Refine research skills through different bacteriological techniques.

+ Analyze and compare the degree of contamination found with past investigations.

+ Raise awareness to the community.

Hypothesis In the past year our fellow classmates were able

to detect the presence of Shigella serotypes A,B and C.

The Caño Martín Peña has excessive levels of coliforms, indicators of fecal matter that represent a hazard for the community.

Hypothesis

+ Because of this information we can say there is a high probability of encountering Salmonella and Shigella SPP. Bacteria due to the high coliform levels resulting from water contamination by fecal matter.

Coliform bacteria Rod-shaped Gram

negative bacillus. Lactose fermenters with

the production of acid and gas when incubated at 35-37°C.

Easy to culture and their presence is used to indicate fecal contamination by other pathogenic organisms in the water.

Enterobacteriaceae Total coliforms:

Escherichia Klebsiella Serratia Citrobacter

Fecal coliform: Eschericia coli

Escherichia coli

Klebsiella pneumoniae

Enterobacter

Serratia marcescens

Citrobacter

Salmonella spp. Vs Shigella spp.

Salmonella spp. Rod-shaped Gram negative

bacillus Non-spore forming. Motile

enterobacteria Facultative anaerobes Hydrogen sulfide

production which can be detected on a TSI growth media

Shigella spp. Rod-shaped Gram

negative bacteria Non-spore forming and

non-motile bacteria

Salmonella Pathogenicity Most of the infections are

due to the ingestion of contaminated food.

Salmonella is responsible for various illnesses such as: Typhoid fever Paratyphoid fever Foodborne illnesses

Enteritis Salmonella also known as food poisoning Salmonella.

Shigella Pathogenicity Most of the infections are

caused via ingestion. The illness is known as

Shigellosis. During infection, Shigella

typically causes dysentery . Invasion of Shigella

Bacterial multiplication Spreading to epithelial cells Tissue destruction

Investigation Site

Station #1 Width : 8 Ft. Depth: 2.05 Ft

Observations Boat nearby Carton Boxes

Station #2 Width: 3 – 5 Ft Depth: 4 Ft

Observations Current of water

coming from a pipe. Fishing traps

Station #3 Width: 8 Ft. Depth: 2 Ft.

Observations Samples were taken

at the shore, because difficult access.

Garbage bags

Methodologies Dilution of sample Membrane filtration Cultivation

SS Agar MacConkey Agar EMB RVS

Identification Enterotubes

Procedure

Dilution Membrane Filtration Cultivation

Dilution of sample

At the Laboratory

Membrane filtration 0.45 μm

At the Laboratory

Membrane filtration

Pure sample Diluted Sample 1:1000

Cultivation

At 37º C. 24 – 48 Hours

Selective MediumsSalmonella

Shigella Agar MacConkey Agar

Selective Mediums

RVS EMB

Taxonomic Determination: Enterotube System

Taxonomic Determination: Enterotube System

Enterotube SystemCitrobacter

freundiiSerratia

plymuthica

Sampling Dates

M1 – 30/ October/ 2012 M2 – 28/ November/ 2012 M3 – 7/ December/ 2012 M4 – 30/ January/ 2013 M5 – 11/ February/ 2013

Types of Bacteria

Characteristics of the Colony

Bacteria

Salmon Colony Klebsiella ozanaePink Colony Enterobacter cloacaePink Colony Klebsiella pneumoniaePink colony Enterobacter aerogenesTransparent Colony ShigellaTransparent Colony YersiniaBlack Colony Citrobacter freundiPale pink Serratia plymuthicaMetallic Green E. coli

Types of BacteriaBacteriaSerratia marcenscensEnterobacter aglomerans

Enterotubes

Shigella Enterotube

FindingsYear Quantity2008 118,0002009 3,335,0002010 6,800,0002011 60,000,0002012 69,000,0002013 25,333,333

Year Quantity2000 4,000,0002001 5,500,0002002 4,000,0002003 2,000,0002004 100,0002005 73,0002006 80,0002007 57,000

Number of bacteria/ ml of water

Findings on Dilutions

Station #1 1 col in 10¯⁷ : 10,000,000 bact/ml

Station #2 4 col in 10⁻⁶: 4,000,000 bact/ml

Station #3 62 col in 10⁻⁶: 62,000,000 bact/ml

Findings on Filtration Station#1

Pure > 100 col (TNTC) Dilute: 5 col * 1000: 5,000 bact/100ml

Station #2 Pure>100 col (TNTC) Dilute: 40 col * 1000: 40,000 bact/100ml

Station #3 Pure>100 col (TNTC) Dilute: 3 col * 1000: 3,000 bact/100ml

Citrobacter freundiCedecea lapageiKlebsiella pneumoniaeYersinia enterocoliticaCitrobacter freundiCedecea lapageiEnterobacter agglomeransKlebsiella ozaenae

Enterobacter agglomerans

Conclusions

The high level of contamination is due to:

Stagnant water in some areas.

Sewage and water discharge in to the cano.

Deposition of debris into the community.

Conclusions of the Research

From 2004 to 2008 the amount of bacteria were less than a million/ ml of water.

This could be due to the removal of debris

Effort of government and people that lives in the community.

+ During our Research period we noticed a sudden decrease in the level of bacteria.

+ Although we were not able to isolate colonies of Salmonella, we did isolate Citrobacter freundi, Yersinia enterocolitica and Shigella Spp.

+ Which are indicative of the severity of contamination and a possible source of contamination with pathogens to the residents of nearby areas.

We recommend: To maintain the movement of dredging and

cleaning of the entire San Juan Estuary. We saw that according to data, levels of bacteria

were down under a million. With a little of effort is possible to maintain to provide a better life to the people.

To Help organizations and people that live there in a constant way not only, to show interest when elections are near.

References Http://www.mayoclinic.com/health/shigella/DSOO

719 Http://www.cdc.gov/salmonella/enteritidis/ Centers for Disease Control and Prevention.

October 2005. Shigellosis. http://www.cdc.gov/nczved/divisions/dfbmd/diseases/shigellosis/

Indian and Northern Affairs Canada. May 2003. First Nations Water Management Strategy. Http://www.ainc-inac.gc.ca/ai/arp/es/0506/fnwms/fnwms-eng.asp

References Public Health Agency of Canada:Notifiable Diseases on-

line. December 2003. Shigellosis. http://dsol-smed.hc-sc.gc.ca/dsol-smed/ndis/diseases/shig_e.html

www.safewater.com World Health Organization. 2005. Guidelines for the

control of shigellosis, including epidemics due to shigella dysenteriae type 1. http://whqlibdoc.who.int/publications/2005/9241592330.pdf

United States Pathogenic Microorganisms and Natural Toxins Handbook. January 2002. Shigella. Http://pdf.usaid.gov/pdf_docs/ PNADO152.pdf